In the field of life science, it is very important to analyze various cellular phenomena such as changes in the amount of cellular calcium, phosphorylation of intracellular proteins, distribution of ATP, which is a source of energy, transcriptional activities of genes and the like. Various types of molecular probes have been created and used for imaging. These imaging technologies are used for observations at various levels from living cells to organisms. For example, labeling and imaging cancer cells enables the evaluation of anticancer agents and visual analysis of cancer metastases. Light probes, in contrast to radioactive probes, which can be used only by qualified researchers in strictly controlled facilities, have attracted attention. Light probes are broadly divided into luminescent probes and light probes, not requiring use in controlled facilities, nor expensive measuring devices. These probes are stable and inexpensive, and are easily handled. Among light probes, fluorescent probes require excitation light, raising difficulties in obtaining information from deep inside individuals where external light sources cannot be used for excitation. Cellular photodamage caused by external lights also presents a problem. On the other hand, luminescent probes are self-luminescent, not requiring excitation light. Various types of probes have been produced and used for imaging at the individual level.
Luciferase probes are the most utilized among luminescent probes, among which beetle luciferases have already been utilized for intracellular imaging and are found to be useful as a visualization probe enabling prolonged imaging (WO 2007/058140, WO 2006/106752). However, the maximum emission wavelength is 535 nm, in contrast with the most suitable wavelengths for imaging individuals: 650-750 nm, which has very low light energy. Cypridina luciferase makes highly useful probes in combination with its luciferin analogues or nano quantum dots, being capable of producing light with a maximum emission wavelength between about 380 nm (near ultraviolet light, Japanese Patent Application No. 2005-169768) and 650 nm (red to near-infrared light with high penetration efficacy at depth in individuals, US 60/907234). It is clear that, particularly, a near-infrared luminescent probe utilizing energy transfer(bioluminescence resonance energy transfer) between a Cypridina luciferase and nano quantum dot conjugate produces light with high permeability in individuals, thereby making a method using such a probe effective. However, the problems of safety and toxicity or the disturbance of life information caused by luminescent probes associated with the imaging of individuals, such as nano quantum dots using biologically toxic metals and not being easily excreted in the liver, and the like, have been pointed out. Moreover, near-infrared luminescent probes comprising a Renilla luciferase and nano quantum dot conjugate have been reported (So M K et al. Nat Biotechnol. 24 (2006) :339-43), raising concerns about safety for the above reasons.
[Patent document 1] WO2007/058140
[Patent document 2] WO2006/106752
[Patent document 3] Japanese Patent Application No. 2005-169768
[Patent document 4] US60/907,234
[Non-patent document 1] So M K et al. Nat. Biotechnol. 2006, 24:339-43